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Influence of foliar ozone injury on root development and root surface fungi of pinto bean plants
INFLUENCE OF FOLIAR OZONE INJURY ON ROOT DEVELOPMENT AND ROOT SURFACE FUNGI OF PINTO BEAN PLANTS W. J. MANNING, W. A. FEDER, P. M. PAPIA (~ I. PERKINS
Department of Environmental Sciences, University of Massachusetts, Waltham, Massachusetts, USA
ABSTRACT
Chronic exposure of Pinto bean plants to levels of ozone sufficient to cause foliar injury adversely affected shoot and root growth and vigour and enhanced senescence. These effects were reflected in quantitative rather than qualitative differences in the successional root surface fungi. More fungal colonies were consistently isolated from the roots and hypocotyls of plants exposed to ozone than from those grown in charcoalfiltered air. Roots and hypocotyls of plants exposed and not exposed to ozone were colonised by the same fungi and each had the same general mycoflora at each sampling period. Rhizobium nodules were found on the roots of plants grown in charcoalfiltered air, but were not found on the roots of plants grown in the ozone chamber.
INTRODUCTION
Plant growth and metabolism exert a stimulatory effect on the soil microflora near plant roots. This microflora in turn affects mineral nutrient availability and may influence the activities of plant pathogens. This unique zone of interactions between plant roots and their associated microflora is called the rhizosphere. Interactions that occur in the rhizosphere can have a pronounced effect on plant growth and soil fertility (Alexander, 1965). Conversely, conditions that significantly affect plant growth and metabolism should also have quantitative and qualitative effects on the rhizosphere microflora (Katzelson, 1965). Environmental factors, such as light, temperature, soil type and reaction, are known to directly affect plant growth and metabolism and may indirectly affect rhizosphere microbial populations in varying degrees (Harley & Waid, 1955a; Peterson, 1958 & 1961; Rouatt et al., 1963; Rovira, 1959; Taylor & Parkinson, 1964). It is also increasingly evident that other environmental factors, 305 Environ. Pollut. (1) (1971) pp, 305-312--O Elsevier Publishing Company Ltd, England--Printed in Great Britain
306
w.J.
MANNING, W. A. FEDER, P. M. PAPIA, I. PERKINS
such as certain levels of ambient air pollution and specific air pollutants, can also directly affect the growth and metabolism of many plants (Engle & Gabelman, 1967; Feder, 1970; Hill et al., 1961; Hindawi, 1968 & 1970; Scheffer & Hedgcock, 1955; Taylor et al., 1958; Tingey et al., 1969). Little is known, however, of the possible influence of the direct effects of air pollutants on plant growth and metabolism in relation to indirect effects on the rhizosphere microflora of plant roots. In this paper we report the results of a study on the influence of tbliar ozone injury on root development and root surface fungi of Pinto bean plants.
MATERIALS AND METHODS
Seed of certified 111 strain of Pinto bean were surface-sterilised for 10 minutes in a solution of 10 ~o chlorine bleach and 2 ~ ethanol, followed by three sterile distilled water rinses. Sound seed were then planted in pots of non-sterile Merrimac fine sandy loam in the greenhouse in June. Half of the pots were placed in a greenhouse fumigation chamber and all emerging plants were exposed to 0.1 to 0.15/~i/1 ozone for eight hours per day for 28 days. Ozone was produced by a Welsbach generator and levels were monitored with Mast meters. Plants used as controls were grown in an ozone-free, charcoal-filtered air greenhouse chamber. All plants were observed daily for ozone injury and general appearance. Plant samples were taken 4, 7, 11, 17, 23, and 28 days after seed was planted. Forty typical plants from each chamber were carefully removed from pots and brought into the laboratory. Roots were washed in running tap water to remove loose soil. Plant height and root length were recorded in cm. Roots and hypocotyls were examined for surface browning, lesions, and decay. Fresh and dry weights in g were obtained for all plants in the final sample. Tissue sections 1 cm in length were excised from the tip, mid-point, and base of the primary root and the mid-point of the below-ground hypocotyl and placed in tubes containing 15 ml of sterile distilled water and shaken vigorously for two minutes. This process was repeated 20 times with changes of wash water after the 1st, 5th, 10th, and 15th washings. Similar root washing techniques have been described (Harley & Waid, 1955b; Parkinson et al., 1963). Following the 20th washing, tissue sections were blotted on sterile filter paper and plated on peptonedextrose agar (PD) containing 10 mg chlortetra-cycline/1. All plates were incubated at 25°C for seven days. Resulting fungal colonies were observed, counted and subcultured for later identification. The taxonomic system of Snyder and Hansen was used to identify Fusaria (Tousson & Nelson, 1968). Steam-sterilised fine sandy loam was infested with vermiculite-potato dextrose broth inoculum (Varney, 1961) of fungi representative of those isolated to determine their pathogenicity to Pinto bean plants in pot tests in the greenhouse. Re-isolations
307
ROOT DEVELOPMENT AND ROOT SURFACE FUNGI OF PINTO BEAN PLANTS RESULTS
Upper surface flecking was observed on the first set of true leaves on plants in the ozone chamber nine days after seed was planted. This effect was intensified within three days to include bronzing and reddening of the flecked areas, marginal burning, yellowing and leaf fall. This progressive syndrome was observed as each set of trifoliate leaves expanded and matured. These results are similar to other descriptions of ozone injury on Pinto bean leaves (Engle & Gabelman, 1967; Hill et al., 1961). Differences in plant height were readily apparent when plants were measured as early as seven days after seed was planted (Table 1). Plants grown in filtered air TABLE 1 COMPARATIVE DEVELOPMENT OF PINTO BEAN PLANTS GROWN IN AN OZONE CHAMBER (O) AND IN AN OZONE-FREE, CARBON-FILTERED AIR CHAMBER (F)
Number of days after planting Measurements Height of tops* (O) (F) Length of roots* (O) (F) Final weightst
(O) (F) *Avg. 40 plants in crn.
4
7
11
17
23
28
4"8 6"0
13"9 19"0
17'7 33-0
22.3 64.0
30"3 65.7
31.7 105.1
5"0 4"0
8"1 9-6
11.4 12-7
9.6 13.4
8.6 26-4
8"1 26"9
Tops
Roots
Fresh Wt Dry Wt
Fresh Wt Dry Wt
38-5 185'7
4"0 24'0
11 "5 45"3
1'0 4'0
tAvg. 40 plants in g.
were two to three times as tall and had leaves and stems darker in colour and twice as large as plants in the ozone chamber. Extensive differences in root length were also observed, starting with plants measured 17 days after seed was planted (Table 1). Root systems of plants from the ozone chamber were markedly decreased in size and vigour and had poorly developed secondary roots. Many root tips were decayed and surface browning and dark lesions were prevalent on all root and hypocotyl regions. At termination, this effect had intensified. Occasional areas of surface browning and necrotic lesions were observed on the roots and hypocotyls of plants from the filtered air chamber. Fresh weight and dry weight differences were very striking (Table 1). These results are similar to other general descriptions of the influence of ambient air pollution and synthetic smog on growth and development of the tops and roots of Pinto bean and other plants (Hindawi, 1968
TABLE 2
163
als
20 38 43 32
(F)
133 22.3
4
atal no. fungal colonies from 40 plant segments.
nere,ase in no. of colonies
29 46 48 40
(O)
of primary root i-section of primary root e of primary root l.section of hypocotyl
ources of washed plant segments used for isolations
159
31 42 40 46
20 37 36 36
(F)
129 23.2
(O)
7
194
44 45 50 55
42 40 44 40
(F)
166 16.8
(O)
11
197
58 46 49 44
36 41 40 42
(F)
159 23.9
(O)
17
207
56 59 50 42
35 48 44 34
(F)
161 28.5
(O)
23
192
48 46 52 46
36 42 40 41
(F)
159 20.7
(O)
28
No. of fungal colonies by days after planting and chamber types*
1112
266 284 289 273
of
increase in no.
907
40.7 11.3 17.0 21.3
(F) colonies 189 246 247 225
22.5
(O)
Totals
SUMMARY OF NUMBERS OF FUNGI ISOLATED FROM HYPOCOTYLS AND ROOT SURFACES OF PINTO BEAN PLANTS GROWN IN AN OZONE CHAMBER (O) AND IN AN OZONE-FREE t CARBON-FILTERED AIR CHAMBER (F)
t~
•
.~
m
.~
Z z
TABLE
3
M
+
+
T*
+
+
4 H
+
+
+ +
B
+
+ +
T
+
+
M
7
+
+
B
+
+
H
+
+ +
+ + +
T
+ +
+
M
11
+
+
B
+
+
+
H
+
+
+ + + +
T
+ +
+
+
M
17
+
+
B
+
+ + +
H
+
+
+
+
T
= tip, M = mid-point, and B = base of the primary root. H = mid-point of below-ground hypocotyl.
;terile white Mycelium
arium oxysporum ~. roseum :. solani ~. moniliforme 'enicillium spp. 'ythium sp. "richoderma lignorum
dominant fungi
+
+ +
+
M
23
+
+ + + +
B
Occurrence o f predominant fungi by no. o f days after planting
+
+ + + +
H
+
+
+
+
T
SUMMARY OF OCCURRENCE OF PREDOMINANT FUNGI ISOLATED FROM HYPOCOTYL A N D ROOT SURFACES OF PINTO BEAN PLANTS G R O W N IN BOTH OZONE A N D FILTERED AIR CHAMBERS
+
+ +
+
M
28
+
+ + + +
B
+
+ + + +
H
tao O
e0
,-] O
O
Z
¢3 t~
Z7
Z
Z
rn m 0
0 o
310
w.J.
MANNING, W. A. FEDER, P. M. PAPIA AND I. PERKINS
roots of plants grown in charcoal-filtered air, but were not found on the roots of plants grown in the ozone chamber. When fungal colonies from plated washed tissues were counted, it was found that more colonies were consistently isolated from the roots and hypocotyls of plants grown in the ozone chamber than from plants grown in the filtered air chamber (Table 2). This was particularly evident for root tips of ozone-injured plants. This effect was apparent before obvious differences in root development and condition were observed (Table 1). While there were quantitative differences in the number of fungal colonies isolated from roots and hypocotyls, there were no significant qualitative differences (Table 3). Roots and hypocotyls of plants from the ozone and filtered air chambers were colonised by the same general mycoflora at each sampling period. Fusarium oxysporum was the predominant fungus isolated from all four types of tissue sections at each sampling period. Other Fusaria became more prevalent from 17-28 days after seed was planted. Other fungi were isolated only from certain root and hypocotyl locations and at certain intervals after seed was planted. A sterile white mycelial form was isolated only from root tips. Penicillium spp. were isolated only during the first three sampling periods. The dominance of the root surface nlycoflora by Fusaria is consistent with the observations of other investigators (Dix, 1964; Parkinson et al., 1963; Peterson, 1958; Taylor & Parkinson, 1965). F. oxysporum was the most frequently isolated fungus from decayed root tips, root and hypocotyl lesions and areas of surface browning. The frequency of isolation of other Fusaria, Trichoderma lignorum, and a Pythium sp. increased as the plants aged. Examination of inoculated Pinto bean plants and PD re-isolation plates showed that 5 0 ~ of the F. oxysporum isolates, 4 0 ~ of the F. solani isolates, 3 5 ~ of the F. roseum isolates, and 60 ~o of the F. moniliforme isolates caused slight surface browning on roots and hypocotyls. With the exception of a few isolates of F. solani, the other isolates of these fungi had no visible effects on Pinto bean roots and hypocotyls. Isolates of T. lignorum, Penicillium spp., and the sterile white mycelial form also had no effects. Several of the Pythium isolates did cause surface browning and lesions on roots and hypocotyls.
DISCUSSION
Foliar ozone injury resulted in reduced over-aU growth and increased senescence in Pinto bean plants. More fungal colonies were isolated from the roots and hypocotyls of injured than uninjured plants indicating definite quantitative differences in root surface mycoflora. Early differences may have been due to altered plant metabolism and increased exudation of nutrients from roots which led to
ROOT DEVELOPMENT AND ROOT SURFACE FUNGI OF PINTO BEAN PLANTS
311
activity of rhizoplane fungi in the primary decomposition process of senescent roots expressed as decayed root tips and root and hypocotyl lesions. Root surface Fusaria and other fungi have been suggested as important saprophytic invaders or weak pathogens of roots of plants in reduced states of vigour or advanced senescence (Dix, 1964; Parkison et al., 1963; Peterson, 1958). Most of the fungi isolated were shown to be non-pathogenic to Pinto bean roots and hypocotyls in pot tests. The rhizoplane has been suggested as being an ideal site to study the specific qualitative effects of plant roots on rhizosphere microorganisms (Clark, 1949). Our results indicate that there were no significant qualitative differences in the successional rhizoplane mycoflora of roots and hypocotyls of plants with and without foliar ozone injury. Similar results have been observed (Peterson, 1961) for the effects of light on the mycoflora of wheat and soybean roots. Chronic exposure of Pinto bean plants to levels of ozone sufficient to injure leaves had adverse effects on shoot and root development and quantitative, rather than qualitative, effects on the successional root surface mycoflora. REFERENCES ALEXANDER, M. (1965). Introduction to soil microbiology. New York, John Wiley & Sons, 472 pp. CLARK, F. E. (1949). Soil micro-organisms and plant growth. Adv. AAron., 1, 241-88. Dlx, N. J. (1964). Colonization and decay of bean roots. Trans. Br. mycol. Soc., 47, 585-92. ENGLE, R. L. & GABELMAN, W. H. (1967). The effects of low levels of ozone on Pinto beans Phaseolus vulgaris L. Prec. Am. Soc. heft. Sci., 91, 304-9. FEDER, W. A. (1970). Plant response to chronic exposure to low levels of oxidant type air pollution. Emqron. Pollut., 1, 73-9. HARLEY, J. L. d~ WAID, J. S. (1955a). The effect of light upon the roots of beech and its surface population. Pl. Soil, 7, 96-112. HARLEY, J. L. d~. WAR), J. S. (1955b). A method of studying active mycelia on living roots and on other surfaces in the soil. Trans. Br mycoL Soc., 38, 104-18. HILL, A. C., PACK, M. R., TRESHOW, M., DOWNS, R. J. • TRANSTRUM, L. G. (1961). Plant injury induced by ozone. Phytopathology, 51, 356-63. HINDAWl, 1. J. (1958). Injury by sulfur dioxide, hydrogen fluoride, and chlorine as observed and reflected on vegetation in the field. J. Air Pollut. Control Ass., 18, 307-12. HINDAWI, I. J. (1970). Air pollution injury to vegetation. National Air Pollution Control Administration Publ. No. AP-71, 44 pp. KATZELSON, H. (1965). Nature and importance of the rhizosphere. In: Ecology of soil-borne plant pathogens, ed. by K. F. Baker & W. C. Snyder. Univ. Calif. Press, Berkeley, 187-209. PARKINSON, D., TAYLOR, G. S. 86 PEARSON, R. (1963). Studies of fungi in the root region. 1. The development of fungi on young roots. PI. Soil, 19, 332-49. PETERSON, E. A. (1958). Observations on fungi associated with plant roots. Can. J. Microbiol., 4, 257-65. PETERSON, E. A. (1961). Observations on the influence of plant illumination on the fungal flora of roots. Can. J. Microbiol., 7, 1-6. ROUATr, J. W., PETERSON, E. A., KATZELSON, H. & HENDERSON, V. E. (1963). Microorganisms in the root zone in relation to temperature. Can. J. Microbiol., 9, 227-36. ROVlRA, A. D. (1959). Root excretions in relation to the rhizosphere effect. IV. Influence of plant species, age of plant, light, temperature, and calcium nutrition on exudation. PI. Soil, 11, 53-64. SCHEFFER, T. C. & HEDGCOCK, G. C. (1955). Injury to Northwestern forest trees by sulfur dioxide from smelters. Tech. Bull. US Dep. Agric. No. 1117, 49 pp. TAYLOR, G. S. & PARKINSON, n . (1964). Studies on fungi in the root region. II. The effects of ~-~;~
~ , , ~ m ~ t o l
~nA;ti~e
~
th~
rl~wolnnm,~nt
n f rc'..~t e t l r f - . I o ~ m v c ' n f l n r a c
t'~f" d w a r f
312
w . J . MANNING, W. A. FEDER, P. M. PAPIA AND I. PERKINS
TAYLOR, G. S. & PARKINSON, D. (1965). Studies on fungi in the root region. IV. Fungi associated with the roots of Phaseolus vulgaris L. PL Soil, 22, 1-20. TAYLOR, O. C., CARDIFF, E. A., MERSEREAU,J. n . & MIDDLETON, J. T. (1958). Effect of air-borne reaction products of ozone and 1-N-hexene vapor (synthetic smog) on growth of avocado seedlings. Proc. Am. Soc. hurt. Sci., 72, 320-5. TINGEY, D. T., HECK, W. W. & REINERT, R. A. (1969). Reduction of radish yields by low concentrations of ozone and sulfur dioxide. Hortscience, 4, 106 (Abstr.). TOUSSON, T. A. & NELSON, P. E. (1968). A pictorial guide to the identification of Fusarium species according to the taxonomic system of Snyder and Hansen. Univ. Park, Penn. State Univ. Press. 51 pp. VARNEY,E. H. (1961). Vermiculite media for growing fungi. PI. Dis. Reptr, 45, 393.
Department of Environmental Sciences, University of Massachusetts, Waltham, Massachusetts, USA
ABSTRACT
Chronic exposure of Pinto bean plants to levels of ozone sufficient to cause foliar injury adversely affected shoot and root growth and vigour and enhanced senescence. These effects were reflected in quantitative rather than qualitative differences in the successional root surface fungi. More fungal colonies were consistently isolated from the roots and hypocotyls of plants exposed to ozone than from those grown in charcoalfiltered air. Roots and hypocotyls of plants exposed and not exposed to ozone were colonised by the same fungi and each had the same general mycoflora at each sampling period. Rhizobium nodules were found on the roots of plants grown in charcoalfiltered air, but were not found on the roots of plants grown in the ozone chamber.
INTRODUCTION
Plant growth and metabolism exert a stimulatory effect on the soil microflora near plant roots. This microflora in turn affects mineral nutrient availability and may influence the activities of plant pathogens. This unique zone of interactions between plant roots and their associated microflora is called the rhizosphere. Interactions that occur in the rhizosphere can have a pronounced effect on plant growth and soil fertility (Alexander, 1965). Conversely, conditions that significantly affect plant growth and metabolism should also have quantitative and qualitative effects on the rhizosphere microflora (Katzelson, 1965). Environmental factors, such as light, temperature, soil type and reaction, are known to directly affect plant growth and metabolism and may indirectly affect rhizosphere microbial populations in varying degrees (Harley & Waid, 1955a; Peterson, 1958 & 1961; Rouatt et al., 1963; Rovira, 1959; Taylor & Parkinson, 1964). It is also increasingly evident that other environmental factors, 305 Environ. Pollut. (1) (1971) pp, 305-312--O Elsevier Publishing Company Ltd, England--Printed in Great Britain
306
w.J.
MANNING, W. A. FEDER, P. M. PAPIA, I. PERKINS
such as certain levels of ambient air pollution and specific air pollutants, can also directly affect the growth and metabolism of many plants (Engle & Gabelman, 1967; Feder, 1970; Hill et al., 1961; Hindawi, 1968 & 1970; Scheffer & Hedgcock, 1955; Taylor et al., 1958; Tingey et al., 1969). Little is known, however, of the possible influence of the direct effects of air pollutants on plant growth and metabolism in relation to indirect effects on the rhizosphere microflora of plant roots. In this paper we report the results of a study on the influence of tbliar ozone injury on root development and root surface fungi of Pinto bean plants.
MATERIALS AND METHODS
Seed of certified 111 strain of Pinto bean were surface-sterilised for 10 minutes in a solution of 10 ~o chlorine bleach and 2 ~ ethanol, followed by three sterile distilled water rinses. Sound seed were then planted in pots of non-sterile Merrimac fine sandy loam in the greenhouse in June. Half of the pots were placed in a greenhouse fumigation chamber and all emerging plants were exposed to 0.1 to 0.15/~i/1 ozone for eight hours per day for 28 days. Ozone was produced by a Welsbach generator and levels were monitored with Mast meters. Plants used as controls were grown in an ozone-free, charcoal-filtered air greenhouse chamber. All plants were observed daily for ozone injury and general appearance. Plant samples were taken 4, 7, 11, 17, 23, and 28 days after seed was planted. Forty typical plants from each chamber were carefully removed from pots and brought into the laboratory. Roots were washed in running tap water to remove loose soil. Plant height and root length were recorded in cm. Roots and hypocotyls were examined for surface browning, lesions, and decay. Fresh and dry weights in g were obtained for all plants in the final sample. Tissue sections 1 cm in length were excised from the tip, mid-point, and base of the primary root and the mid-point of the below-ground hypocotyl and placed in tubes containing 15 ml of sterile distilled water and shaken vigorously for two minutes. This process was repeated 20 times with changes of wash water after the 1st, 5th, 10th, and 15th washings. Similar root washing techniques have been described (Harley & Waid, 1955b; Parkinson et al., 1963). Following the 20th washing, tissue sections were blotted on sterile filter paper and plated on peptonedextrose agar (PD) containing 10 mg chlortetra-cycline/1. All plates were incubated at 25°C for seven days. Resulting fungal colonies were observed, counted and subcultured for later identification. The taxonomic system of Snyder and Hansen was used to identify Fusaria (Tousson & Nelson, 1968). Steam-sterilised fine sandy loam was infested with vermiculite-potato dextrose broth inoculum (Varney, 1961) of fungi representative of those isolated to determine their pathogenicity to Pinto bean plants in pot tests in the greenhouse. Re-isolations
307
ROOT DEVELOPMENT AND ROOT SURFACE FUNGI OF PINTO BEAN PLANTS RESULTS
Upper surface flecking was observed on the first set of true leaves on plants in the ozone chamber nine days after seed was planted. This effect was intensified within three days to include bronzing and reddening of the flecked areas, marginal burning, yellowing and leaf fall. This progressive syndrome was observed as each set of trifoliate leaves expanded and matured. These results are similar to other descriptions of ozone injury on Pinto bean leaves (Engle & Gabelman, 1967; Hill et al., 1961). Differences in plant height were readily apparent when plants were measured as early as seven days after seed was planted (Table 1). Plants grown in filtered air TABLE 1 COMPARATIVE DEVELOPMENT OF PINTO BEAN PLANTS GROWN IN AN OZONE CHAMBER (O) AND IN AN OZONE-FREE, CARBON-FILTERED AIR CHAMBER (F)
Number of days after planting Measurements Height of tops* (O) (F) Length of roots* (O) (F) Final weightst
(O) (F) *Avg. 40 plants in crn.
4
7
11
17
23
28
4"8 6"0
13"9 19"0
17'7 33-0
22.3 64.0
30"3 65.7
31.7 105.1
5"0 4"0
8"1 9-6
11.4 12-7
9.6 13.4
8.6 26-4
8"1 26"9
Tops
Roots
Fresh Wt Dry Wt
Fresh Wt Dry Wt
38-5 185'7
4"0 24'0
11 "5 45"3
1'0 4'0
tAvg. 40 plants in g.
were two to three times as tall and had leaves and stems darker in colour and twice as large as plants in the ozone chamber. Extensive differences in root length were also observed, starting with plants measured 17 days after seed was planted (Table 1). Root systems of plants from the ozone chamber were markedly decreased in size and vigour and had poorly developed secondary roots. Many root tips were decayed and surface browning and dark lesions were prevalent on all root and hypocotyl regions. At termination, this effect had intensified. Occasional areas of surface browning and necrotic lesions were observed on the roots and hypocotyls of plants from the filtered air chamber. Fresh weight and dry weight differences were very striking (Table 1). These results are similar to other general descriptions of the influence of ambient air pollution and synthetic smog on growth and development of the tops and roots of Pinto bean and other plants (Hindawi, 1968
TABLE 2
163
als
20 38 43 32
(F)
133 22.3
4
atal no. fungal colonies from 40 plant segments.
nere,ase in no. of colonies
29 46 48 40
(O)
of primary root i-section of primary root e of primary root l.section of hypocotyl
ources of washed plant segments used for isolations
159
31 42 40 46
20 37 36 36
(F)
129 23.2
(O)
7
194
44 45 50 55
42 40 44 40
(F)
166 16.8
(O)
11
197
58 46 49 44
36 41 40 42
(F)
159 23.9
(O)
17
207
56 59 50 42
35 48 44 34
(F)
161 28.5
(O)
23
192
48 46 52 46
36 42 40 41
(F)
159 20.7
(O)
28
No. of fungal colonies by days after planting and chamber types*
1112
266 284 289 273
of
increase in no.
907
40.7 11.3 17.0 21.3
(F) colonies 189 246 247 225
22.5
(O)
Totals
SUMMARY OF NUMBERS OF FUNGI ISOLATED FROM HYPOCOTYLS AND ROOT SURFACES OF PINTO BEAN PLANTS GROWN IN AN OZONE CHAMBER (O) AND IN AN OZONE-FREE t CARBON-FILTERED AIR CHAMBER (F)
t~
•
.~
m
.~
Z z
TABLE
3
M
+
+
T*
+
+
4 H
+
+
+ +
B
+
+ +
T
+
+
M
7
+
+
B
+
+
H
+
+ +
+ + +
T
+ +
+
M
11
+
+
B
+
+
+
H
+
+
+ + + +
T
+ +
+
+
M
17
+
+
B
+
+ + +
H
+
+
+
+
T
= tip, M = mid-point, and B = base of the primary root. H = mid-point of below-ground hypocotyl.
;terile white Mycelium
arium oxysporum ~. roseum :. solani ~. moniliforme 'enicillium spp. 'ythium sp. "richoderma lignorum
dominant fungi
+
+ +
+
M
23
+
+ + + +
B
Occurrence o f predominant fungi by no. o f days after planting
+
+ + + +
H
+
+
+
+
T
SUMMARY OF OCCURRENCE OF PREDOMINANT FUNGI ISOLATED FROM HYPOCOTYL A N D ROOT SURFACES OF PINTO BEAN PLANTS G R O W N IN BOTH OZONE A N D FILTERED AIR CHAMBERS
+
+ +
+
M
28
+
+ + + +
B
+
+ + + +
H
tao O
e0
,-] O
O
Z
¢3 t~
Z7
Z
Z
rn m 0
0 o
310
w.J.
MANNING, W. A. FEDER, P. M. PAPIA AND I. PERKINS
roots of plants grown in charcoal-filtered air, but were not found on the roots of plants grown in the ozone chamber. When fungal colonies from plated washed tissues were counted, it was found that more colonies were consistently isolated from the roots and hypocotyls of plants grown in the ozone chamber than from plants grown in the filtered air chamber (Table 2). This was particularly evident for root tips of ozone-injured plants. This effect was apparent before obvious differences in root development and condition were observed (Table 1). While there were quantitative differences in the number of fungal colonies isolated from roots and hypocotyls, there were no significant qualitative differences (Table 3). Roots and hypocotyls of plants from the ozone and filtered air chambers were colonised by the same general mycoflora at each sampling period. Fusarium oxysporum was the predominant fungus isolated from all four types of tissue sections at each sampling period. Other Fusaria became more prevalent from 17-28 days after seed was planted. Other fungi were isolated only from certain root and hypocotyl locations and at certain intervals after seed was planted. A sterile white mycelial form was isolated only from root tips. Penicillium spp. were isolated only during the first three sampling periods. The dominance of the root surface nlycoflora by Fusaria is consistent with the observations of other investigators (Dix, 1964; Parkinson et al., 1963; Peterson, 1958; Taylor & Parkinson, 1965). F. oxysporum was the most frequently isolated fungus from decayed root tips, root and hypocotyl lesions and areas of surface browning. The frequency of isolation of other Fusaria, Trichoderma lignorum, and a Pythium sp. increased as the plants aged. Examination of inoculated Pinto bean plants and PD re-isolation plates showed that 5 0 ~ of the F. oxysporum isolates, 4 0 ~ of the F. solani isolates, 3 5 ~ of the F. roseum isolates, and 60 ~o of the F. moniliforme isolates caused slight surface browning on roots and hypocotyls. With the exception of a few isolates of F. solani, the other isolates of these fungi had no visible effects on Pinto bean roots and hypocotyls. Isolates of T. lignorum, Penicillium spp., and the sterile white mycelial form also had no effects. Several of the Pythium isolates did cause surface browning and lesions on roots and hypocotyls.
DISCUSSION
Foliar ozone injury resulted in reduced over-aU growth and increased senescence in Pinto bean plants. More fungal colonies were isolated from the roots and hypocotyls of injured than uninjured plants indicating definite quantitative differences in root surface mycoflora. Early differences may have been due to altered plant metabolism and increased exudation of nutrients from roots which led to
ROOT DEVELOPMENT AND ROOT SURFACE FUNGI OF PINTO BEAN PLANTS
311
activity of rhizoplane fungi in the primary decomposition process of senescent roots expressed as decayed root tips and root and hypocotyl lesions. Root surface Fusaria and other fungi have been suggested as important saprophytic invaders or weak pathogens of roots of plants in reduced states of vigour or advanced senescence (Dix, 1964; Parkison et al., 1963; Peterson, 1958). Most of the fungi isolated were shown to be non-pathogenic to Pinto bean roots and hypocotyls in pot tests. The rhizoplane has been suggested as being an ideal site to study the specific qualitative effects of plant roots on rhizosphere microorganisms (Clark, 1949). Our results indicate that there were no significant qualitative differences in the successional rhizoplane mycoflora of roots and hypocotyls of plants with and without foliar ozone injury. Similar results have been observed (Peterson, 1961) for the effects of light on the mycoflora of wheat and soybean roots. Chronic exposure of Pinto bean plants to levels of ozone sufficient to injure leaves had adverse effects on shoot and root development and quantitative, rather than qualitative, effects on the successional root surface mycoflora. REFERENCES ALEXANDER, M. (1965). Introduction to soil microbiology. New York, John Wiley & Sons, 472 pp. CLARK, F. E. (1949). Soil micro-organisms and plant growth. Adv. AAron., 1, 241-88. Dlx, N. J. (1964). Colonization and decay of bean roots. Trans. Br. mycol. Soc., 47, 585-92. ENGLE, R. L. & GABELMAN, W. H. (1967). The effects of low levels of ozone on Pinto beans Phaseolus vulgaris L. Prec. Am. Soc. heft. Sci., 91, 304-9. FEDER, W. A. (1970). Plant response to chronic exposure to low levels of oxidant type air pollution. Emqron. Pollut., 1, 73-9. HARLEY, J. L. d~ WAID, J. S. (1955a). The effect of light upon the roots of beech and its surface population. Pl. Soil, 7, 96-112. HARLEY, J. L. d~. WAR), J. S. (1955b). A method of studying active mycelia on living roots and on other surfaces in the soil. Trans. Br mycoL Soc., 38, 104-18. HILL, A. C., PACK, M. R., TRESHOW, M., DOWNS, R. J. • TRANSTRUM, L. G. (1961). Plant injury induced by ozone. Phytopathology, 51, 356-63. HINDAWl, 1. J. (1958). Injury by sulfur dioxide, hydrogen fluoride, and chlorine as observed and reflected on vegetation in the field. J. Air Pollut. Control Ass., 18, 307-12. HINDAWI, I. J. (1970). Air pollution injury to vegetation. National Air Pollution Control Administration Publ. No. AP-71, 44 pp. KATZELSON, H. (1965). Nature and importance of the rhizosphere. In: Ecology of soil-borne plant pathogens, ed. by K. F. Baker & W. C. Snyder. Univ. Calif. Press, Berkeley, 187-209. PARKINSON, D., TAYLOR, G. S. 86 PEARSON, R. (1963). Studies of fungi in the root region. 1. The development of fungi on young roots. PI. Soil, 19, 332-49. PETERSON, E. A. (1958). Observations on fungi associated with plant roots. Can. J. Microbiol., 4, 257-65. PETERSON, E. A. (1961). Observations on the influence of plant illumination on the fungal flora of roots. Can. J. Microbiol., 7, 1-6. ROUATr, J. W., PETERSON, E. A., KATZELSON, H. & HENDERSON, V. E. (1963). Microorganisms in the root zone in relation to temperature. Can. J. Microbiol., 9, 227-36. ROVlRA, A. D. (1959). Root excretions in relation to the rhizosphere effect. IV. Influence of plant species, age of plant, light, temperature, and calcium nutrition on exudation. PI. Soil, 11, 53-64. SCHEFFER, T. C. & HEDGCOCK, G. C. (1955). Injury to Northwestern forest trees by sulfur dioxide from smelters. Tech. Bull. US Dep. Agric. No. 1117, 49 pp. TAYLOR, G. S. & PARKINSON, n . (1964). Studies on fungi in the root region. II. The effects of ~-~;~
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TAYLOR, G. S. & PARKINSON, D. (1965). Studies on fungi in the root region. IV. Fungi associated with the roots of Phaseolus vulgaris L. PL Soil, 22, 1-20. TAYLOR, O. C., CARDIFF, E. A., MERSEREAU,J. n . & MIDDLETON, J. T. (1958). Effect of air-borne reaction products of ozone and 1-N-hexene vapor (synthetic smog) on growth of avocado seedlings. Proc. Am. Soc. hurt. Sci., 72, 320-5. TINGEY, D. T., HECK, W. W. & REINERT, R. A. (1969). Reduction of radish yields by low concentrations of ozone and sulfur dioxide. Hortscience, 4, 106 (Abstr.). TOUSSON, T. A. & NELSON, P. E. (1968). A pictorial guide to the identification of Fusarium species according to the taxonomic system of Snyder and Hansen. Univ. Park, Penn. State Univ. Press. 51 pp. VARNEY,E. H. (1961). Vermiculite media for growing fungi. PI. Dis. Reptr, 45, 393.